基于Na+通道的“开/关”状态研究铁棒锤中乌头碱型生物碱砂炒减毒的关键化学基团

Different with boiling or steaming methods, the pharmacist of Zang and Qiang nationality used sand frying method to process Aconitum pendulum. We have demonstrated that during the procedure of sand frying, aconitine and deoxyaconitine convert into 16-epi-pyroaconitine and 16-epi-pyrodeoxyaconitine, respectively. However, the structure conversion pattern of aconitine type diterpenoid alkaloids and the key group of pyrolysis products associated with cardiotoxicity are still unknown. Inhibition of the voltage-gated Na+ channel inactivation is an important mechanism of cardiotoxicity induced by aconitine type diterpenoid alkaloids. Therefore, we hypothesized that the conversion of certain groups within the aconitine type diterpenoid alkaloids, resulting in the inhibition of Na+ channel inactivation effect disappeared..In the present study, we use a variety of chromatographic and spectral methods for isolation and identification of compounds from sand fried Aconitum pendulum. Whole cell patch clamp technique will be used to compare the effects of aconitine type diterpenoid alkaloids and their pyrolysis products on Na+ channel, and animal model of arrhythmia will be used to evaluate their cardiotoxicity. The results of our research is a beneficial exploration for the science of processing of traditional Chinese Medicine.

藏医、羌医通常采用砂炒法炮制有毒药物铁棒锤。课题组发现，与水煮和蒸制法炮制原理不同，砂炒过程中乌头碱、去氧乌头碱通过热解方式转化为16-epi-pyroaconitine和16-epi-pyrodeoxyaconitine，炮制前后有多个位置的取代基发生转化，但转化的基团与心脏毒性的相关性尚不清楚。抑制Na+通道失活是乌头碱型生物碱诱发心律失常（心脏毒性）的电生理机制。我们推测：随着某些取代基在砂炒过程中发生转化，原型生物碱抑制Na+通道失活的作用随之消失，从而实现减毒目的。.因此，本研究：1.拟对铁棒锤中乌头碱型生物碱的砂炒转化成分进行定向分离，明确砂炒过程中其结构转化的完整方式；2.采用膜片钳技术比较原型生物碱与转化成分对Na+通道的影响，并结合整体动物实验，明确与心脏毒性相关的关键取代基。研究成果能够为揭示铁棒锤砂炒法的科学性提供依据，对于乌头类药物的炮制研究也是一种有益探索。

本研究采用油浴的方式模拟砂炒过程，筛选出乌头碱砂炒转化的温度参数为140℃～160℃，时间参数为30～40 min。3-乙酰乌头碱砂炒转化的温度参数为160℃～180℃，时间参数为20～40min。通过对砂炒产物的分离鉴定，阐明乌头碱、3-乙酰乌头碱的转化途径：砂炒过程中乌头碱依次转化为pyroaconitine和16-epi-pyroaconitine；3-乙酰乌头碱依次转化为pyro-3-acetylaconitine和16-epi-pyro-3-acetylaconitine。另外，本研究从康定乌头中分离得到与乌头碱结构相似的滇乌头碱，能够为下一步乌头碱类生物碱砂炒炮制原理研究奠定基础。